Mannitol-1-phosphate dehydrogenase activity in Ectocarpus siliculosus, a key role for mannitol synthesis in brown algae

Abstract

Mannitol represents a major end product of photosynthesis in brown algae (Phaeophyceae), and is, with the β-1,3-glucan laminarin, the main form of carbon storage for these organisms. Despite its importance, little is known about the genes and enzymes responsible for the metabolism of mannitol in these seaweeds. Taking benefit of the sequencing of the Ectocarpus siliculosus genome, we focussed our attention on the first step of the synthesis of mannitol (reduction of the photo-assimilate fructose-6-phosphate), catalysed by the mannitol-1-phosphate dehydrogenase (M1PDH). This activity was measured in algal extracts, and was shown to be regulated by NaCl concentration in the reaction medium. Genomic analysis revealed the presence of three putative M1PDH genes (named EsM1PHD1, EsM1PDH2 and EsM1PDH3). Sequence comparison with orthologs demonstrates the modular architecture of EsM1PHD1 and EsM1PDH2, with an additional N-terminal domain of unknown function. In addition, gene expression experiments carried out on samples harvested through the diurnal cycle, and after several short-term saline and oxidative stress treatments, showed that EsM1PDH1 is the most highly expressed of these genes, whatever the conditions tested. In order to assess the activity of the corresponding protein, this gene was expressed in Escherichia coli. Cell-free extracts prepared from bacteria containing EsM1PDH1 displayed higher M1PDH activity than bacteria transformed with an empty plasmid. Further characterisation of recombinant EsM1PDH1 activity revealed its very narrow substrate specificity, salt regulation, and sensitivity towards an inhibitor of SH-enzymes.

Effects of hypo- and hypersalinity on photosynthetic performance of Sargassum fusiforme (Fucales, Heterokontophyta)

“…The immersion treatment in 90‰ seawater lasted only 2 h in total in our present work, thus before achieving new homeostasis by synthesis of osmolytes, e.g., mannitol (Reed et al 1985, Rousvoal et al 2011, which requires from several hours to 23 days, dehydration of S. fusiforme cells was inevitable during HL stress. However, it seems not likely that the inhibition of xanthophyll cycle was resulted from dehydration, since S. fusiforme could recover from 6 h of desiccation in air (Pang et al 2007), and xanthophyll cycle rarely ceased during desiccation in most desiccation-tolerant algae (Fernández-Marín et al 2009, Fernández-Marín et al 2011.…”

Mannitol in six autotrophic stramenopiles and Micromonas

“…Aas, 2 Berit Smestad Paulsen, 2 Catherine Boyen, 3,4 Bente Edvardsen 1 an M1P phosphatase (M1Pase); (3) the oxidation of mannitol via the activity of a mannitol-2-dehydrogenase (M2DH) yielding fructose; and (4) the phosphorylation of fructose yielding F6P and involving a hexokinase (HK). 1,2 The first completed draft of a brown algal genome enabled the identification of candidate genes for each of these steps. 3 As these genes were not found in the genomes of the diatoms Thalassiosira pseudonana and Phaeodactylum tricornutum, mannitol metabolism in stramenopiles was considered a trait typical for brown algae.…”

Toward Systems Biology in Brown Algae to Explore Acclimation and Adaptation to the Shore Environment

“…In addition, systems biology approach should allow generating some hypothesis, which could be tested by combining targeted analyses at the biochemical and genetic level to go deeper in the understanding of the mechanisms of stress response. There are a few examples of functional characterization of enzymes from Ectocarpus: GSTs (de Franco et al, 2009), mannitol-1-phosphate dehydrogenase (Rousvoal et al, 2011), GDP-mannose dehydrogenase (Tenhaken et al, 2011). In parallel, forward and reverse genetics approaches are needed to characterize some of the molecular actors involved in abiotic stress reponse.…”

“…Other metabolic reconstructions have been performed following similar trends with great successes, for instance, in several land plants (Dal'Molin et al, 2010aSaha et al, 2011;UrbanczykWochniak and Sumner, 2007;Zhang et al, 2010). In the context of brown algae, the mannitol metabolism, which has a central role in the physiology of these organisms and for which molecular data have been recently published (Michel et al, 2010a;Rousvoal et al, 2011), should be considered as the core biological information to use for anchoring the Ectocarpus metabolic network. Building a network centered on these reactions is a way to attest that the metabolism, as reconstructed, will be appropriate to analyze phenotypical changes of brown algae under different abiotic treatments.…”